DEG_Kelch_Keap1_2

Cellular response against oxidative or electrophilic attack is regulated by the action of a cytoprotective mechanism that functions via the KEAP1 (Q14145)-Nrf2 (Q16236) pathway. This mechanism is conserved from fly to human. The turnover of Nrf2 is maintained by the E3 ligase KEAP1-mediated proteasomal degradation via a two-site substrate recognition mechanism, in which two kelch domains (PF01344) of KEAP1 homodimer associate with one molecule of Nrf2 by a hinge and latch mechanism. The Kelch beta-propeller domain in KEAP1 is a protein hub in oxidative stress response pathway and it recognizes high and low affinity degron motifs in its binding partners. KEAP1-mediated substrate recognition is important for the elimination of its substrate over-activation or otherwise it may be the cause of diseases such as cancer.

This is the second and weaker binding motif present in targets of KEAP1 (Q14145) such as Nrf2 (Q60795). This motif works to correctly position the lysines within the Nrf2 Neh2 domain for efficient ubiquitylation. As in the case of the high affinity motif, this low affinity motif also binds to the bottom face of the β-propeller structure of Kelch domain (PF01344) but with fewer electrostatic interactions and also the motif is partially entrenched, which makes the interaction much weaker. The low affinity motifs are more prone to changes in its binding conformation in response to electrophilic stress, which reflects its important role in the activation of Nrf2.

Oxidative and xenobiotic stresses affect the cellular homeostasis and are responsible for the development of many diseases, including cancer, cardiovascular disease, diabetes, and neurodegeneration. The cellular defence mechanism against the oxidative stress and electrophilic attack referred to as “cytoprotective response” is obtained by the action of an array of genes encoding detoxifying and anti-oxidative stress enzymes. A Cap N’ Collar (CNC) family transcription factor Nrf2 and its homologues act as major regulators of cytoprotective gene expression (Kobayashi,2002). The level of Nrf2 (Q16236) in the cell is modulated mainly by its interaction with the KEAP1 protein (Q14145). One Nrf2 molecule interacts with two molecules of KEAP1 via two binding sites, the stronger binding motif and the weaker binding motif. Under unstressed conditions, KEAP1 Kelch domain (PF01344) binds to the degron motifs in Nrf2 and promote its ubiquitin dependent degradation. When cells are exposed to oxidative stress, the conformational changes in KEAP1 allow the high affinity motif to be attached to the KEAP1, acting as a hinge of the KEAP1 repression gate. On the other hand, the DLG motif acts as a latch, resulting in the detachment of low affinity motif from KEAP1 and escape protein degradation driving the expression of genes encoding detoxifying and anti-oxidative stress enzymes (24024136). KEAP1 is a crucial hub in the oxidative stress responsive pathway and apoptosis (Hast,2013). It acts as a substrate adaptor protein for a Cul3 (Q13618)-dependent E3 ubiquitin ligase. Many of its interacting partners compete with Nrf2 for KEAP1 interaction. They all bind to the same bottom region of the propeller domain of KEAP1 with varying affinities (Padmanabhan,2006). Although there is no obvious sequence similarity between the high and low affinity binding motifs, the residues that are largely buried in the Kelch domain binding interface all have similar φ and ψ dihedral angles. The high affinity motif is present in a higher number of proteins. The low affinity motif, which have been so far identified in only Nrf2 (Q60795) and apoptotic protein Bcl2 (P10417).